Apparatus And Method For Handling Fluids For Analysis

ABSTRACT

A reaction vessel with a bottom drain opening supporting a selected unpressured head of fluid by the surface tension of the fluid. A device processing zone includes a support for spaced rows of reaction vessels, passages communicating with their drain openings of supported vessels, and a pressure source for selectively draining fluid through the drain openings. Generally horizontal bar magnets are supported for selected vertical movement between the vessel rows. A dispensing head has X discharge openings selectively positionable over X selected reaction vessels. A metering pump mechanism selectively meters X a selected quantity of fluid a bulk supply (where X is at least four), and selectively pumps the metered selected quantities through the drain openings to the selected reaction vessels. Methods of drawing fluid from the vessels using the pressure source, and moving the magnets to form a pellet of analyte are also included.

CROSS REFERENCE TO RELATED APPLICATION(S)

This application is a divisional application claiming priority from U.S.patent application Ser. No. 10/865,207, filed Jun. 10, 2004, whichclaims priority from U.S. Provisional Application Ser. No. 60/479,710,filed Jun. 19, 2003.

TECHNICAL FIELD

The present invention is directed toward testing of specimens, andparticularly toward an apparatus and method for processing specimensduring testing, including adding fluids such as reagents during theprocessing of specimens.

BACKGROUND OF THE INVENTION AND TECHNICAL PROBLEMS POSED BY THE PRIORART

Testing sample biological specimens is commonly done, for example, tocheck for the presence of an item of interest, which item may be orinclude all or portions of a specific region of DNA, RNA, or fragmentsthereof, complements, peptides, polypeptides, enzymes, prions, proteins,messenger RNA, transfer RNA, mitochondrial RNA or DNA, antibodies,antigens, allergens, parts of biological entities such as cells, vironsor the like, surface proteins, or functional equivalents of the above,etc. Specimens such as a patient's body fluids (e.g., serum, wholeblood, urine, swabs, plasma, cerebral-spinal fluid, lymph fluids, tissuesolids) can be analyzed using a number of different tests to provideinformation about a patient's health.

In such testing, it is imperative that the specimens be handled in amanner which prevents contaminants from being introduced to thespecimens, whether from the outside environment or between specimens.For example, where the HIV virus from one specimen is inadvertentlyallowed to contaminate the specimen of a different patient, theresulting false positive test result could potentially have catastrophicpsychological effect on the patient, even should subsequent testinglater discover the error. Moreover, since such testing is highlysensitive, even the smallest amounts of contamination can causeerroneous test results. Simply put, it is imperative that the specimensbe properly handled.

In such sophisticated testing, it is also imperative that the variousreagents which may be used in the testing be properly handled as well,not only to avoid contaminants but also to ensure that the properreagent in proper quantities is used at appropriate times.

Commonly, such testing is accomplished using automated devices whichhandle multiple specimens and fluids (typically, reagents). Suchautomated devices typically will use sets of pipettes to move variousfluids between their original containers (usually receptacles such asopen topped tubes) and containers in which the specimens are to beprocessed. For example, a set of 8 specimens may be contained in 8 tubesor other receptacles loaded in a rack on the device, and a head carrying8 pipettes will, through programmed motion, move the pipettes into those8 tubes, where a vacuum will be applied to extract a selected amount ofeach specimen from its tube into the pipettes. The head will thenretract the pipettes from the tubes and move to another set of tubeslocated at a processing station, depositing the extracted amounts ofeach specimen from the pipettes into sets of testing tubes.

In such automated devices, racks or trays of multiple tubes are usuallymoved from one station to the next for different stages of processing.For example, a heating element may be provided at one station, and amagnetic element introducing a magnetic field in the tubes may beprovided at another station. Further, in such situations, multiple traysof multiple tubes may be actively processed in series and simultaneouslyat different stations. However, such processing can result in resourcecontention, where one rack of tubes is delayed from being placed at astation while another rack of tubes completes its processing at thatstation when, as is commonly the case, the processing time at onestation is different than the processing time at another station. Forexample, a tray of tubes which have completed processing at one stationmay be delayed from being processed at the next station until anothertray of tubes at that next station has completed its processing there.Like a chain which is no stronger than its weakest link, such anautomated device will provide an overall processing time which is afunction of the processing time at the slowest station. Of course, aslow overall processing time can reduce the amount of tests which areperformed during a given day, and thereby either delay the completion oftests or require significant additional investment of capital foradditional devices to allow for a desired testing capacity level.

At the processing stations of such automated devices, the specimens arevariously handled according to the purpose of the testing (e.g.,incubated, prepared, lysed, eluted, etc.). For example, the specimensmay be prepared for analyzing, as for example by separating DNA or RNAfrom the specimen. The specimens may also or alternatively be analyzed.Commonly, such processes involve the addition of various fluids(typically reagents) to the specimen in each tube. For example, in afirst step, a reagent may be added to each of the tubes to wash thespecimens, and second and third (and more) reagents may be added to thespecimens in the course of carrying out other processes to, for example,unbind and/or separate the DNA or RNA of interest so that it may beextracted from the specimen in each tube for subsequent testing. Similarprocesses, in which the same or different reagents are added to and/orextracted from the tubes, may also occur after the specimen has beenprepared as part of analyzing the prepared specimens.

In some processes, magnetic fields have been used to assist inseparating analytes of interest from the fluid in the tubes. Forexample, analytes of interest have been bound to magnetic particleswithin a reagent a magnetic field applied to pull the particles andbound analyte to one side of the tube, whereby the reagent may be drawnout of the tube to leave a concentration of the analyte therein. Whereit has been necessary to adjust the magnetic field within the tubes(e.g., in order to change the location where the analytes are to bedrawn), the tubes have been moved in order to accomplish the desiredorientation of the magnetic field in the tubes.

The handling of the reagents and other fluids with automated devicessuch as described above can be problematic. Though the reagents may beautomatically moved from receptacles to the specimen containing tubes inthe processing station by use of the head and pipettes such as noted, itis in the first instance necessary to load the appropriate reagent intothe appropriate receptacle on the device in order to ensure that thehead and pipettes are adding the appropriate reagent to the appropriatespecimen containing tube at the appropriate time in the process.Further, it should be recognized that it is necessary for thereceptacles to be readily cleaned, whether to remove possiblecontaminants or to permit use of different fluids in connection withdifferent processes. As a result of such requirements, the receptaclesare typically readily removable from the apparatus for such action.

Heretofore, loading the appropriate reagent into the appropriatereceptacle has been accomplished in several different ways. In one suchprocedure, the individual who is controlling the device manuallymeasures and adds the reagents to receptacles, and then places thosereceptacles on the device. In another such procedure, the loading ofreagents is automatically accomplished by the device itself, which usessome transfer apparatus (such as a head and pipette(s) as previouslydescribed) to move the reagents from bulk supplies of the reagentsprovided with the device.

Removing reagents from tubes is similarly accomplished, such as by useof a head which positions pipettes in the tube and vacuum draws thefluid from the tubes into the pipettes. Such a process can be timeconsuming, and tie up the head from other uses, particularly ifprevention of contamination between tubes makes it necessary to use anew pipette with each tube. In such cases, it may be necessary torepetitively move the head to discharge, discard and pick up newpipettes every time fluid is drawn from tubes (e.g., a head carryingeight pipettes may have to be cycled six times when used with a tray of48 tubes, where each cycle requires discharging and discarding usedpipettes, and picking up new pipettes). Of course, in such situations,multiple pipettes will be consumed at some cost. U.S. Pat. No. 6,117,398alternatively discloses drawing fluid from the bottom of a samplevessel, wherein a valve is situated between every sample processingvessel and the waste container.

The present invention is directed to overcoming one or more of theproblems as set forth above.

SUMMARY OF THE INVENTION

In one aspect of the present invention, a reaction vessel for testing ananalyte in a fluid is provided. The vessel has an open top and a drainopening in its bottom, with the drain opening being adapted to support aselected head of fluid and to drain fluid therethrough when a selectedpressure differential exists between the top of the fluid and the bottomof the vessel.

In one form of this aspect of the invention, the surface tension of thefluid supports the selected head of fluid when the pressure differentialbetween the top of the fluid and the bottom of the vessel is less thanthe selected pressure differential.

In another form of this aspect of the invention, a hydrophobic frit isassociated with the drain opening.

In another form of this aspect of the invention, the drain openingpermits draining of the fluid only when the relative pressure betweenthe top of the fluid and the drain opening is at least a selectedamount.

In still another form, a non-wettable surface is provided around thedrain opening on the outside of the vessel.

In yet another form, a drain opening protrusion extends beyond thebottom surface of the vessel.

In another aspect of the present invention, a processing zone for aspecimen handling device is provided, including a support for aplurality of reaction vessels having drain openings in their bottoms,passages adapted to communicate with the bottom drain openings ofsupported reaction vessels, and a source of air at non-atmosphericpressure adapted to selectively drain fluid through the drain openingsin supported reaction vessels. The drain openings are adapted to supporta selected head of fluid in the vessels.

In one form of this aspect of the invention, the source of air atnon-atmospheric pressure is a vacuum source for drawing a vacuum in thepassages.

In another form of this aspect of the invention, a heater is providedfor heating reaction vessels supported in the processing zone.

In still another form, the reaction vessels are adapted to selectivelycontain fluids having a surface tension sufficient to support a selectedhead of fluid without the fluid draining through the drain openings, andthe source of air at non-atmospheric pressure is adapted to selectivelycreate a relatively lower pressure at the drain opening than at the topof the fluid to overcome the fluid surface tension and selectively drainthe fluid through the drain openings.

In yet another form, the support is adapted to support the plurality ofreaction vessels in at least two rows, each row having a defined spacefrom at least one adjacent row. In a further form, the defined spacebetween rows is a generally vertical longitudinal slot, and a bar magnetextends generally horizontally and is supported for selected verticalmovement in the slot. In a still further form, the support is adapted tosupport the plurality of reaction vessels in at least four rows and thedefined space is a generally vertical longitudinal first slot betweenthe first and second rows and a generally vertical longitudinal secondslot between the third and fourth rows, with the bar magnet being afirst bar magnet supported for selected vertical movement in the firstslot and a second bar magnet supported for selected vertical movement inthe second slot. In a yet further form, the first and second bar magnetsare supported for vertical movement together.

In still another aspect of the present invention, a method of processinganalytes in fluids in reaction vessels is provided, including the stepsof (1) supporting a reaction vessel containing an analyte in a fluid,the reaction vessel having a drain opening in its bottom capable ofsupporting a selected head of the fluid in the vessel by the surfacetension of the fluid, (2) drawing the analyte to a side of the vessel toconcentrate the analyte clear of the drain opening, and (3) selectivelycreating a pressure differential between the top of the fluid and thebottom of the drain opening sufficient to overcome the fluid surfacetension and drain the fluid through the drain opening.

In one form of this aspect of the invention, the selectively creating apressure differential step includes selectively creating a vacuumbeneath the drain opening.

In another form of this aspect of the invention, the drawing stepincludes binding the analyte to a magnetic particle, and introducing amagnetic field into the vessel which draws the magnetic particle andbound analyte to the side of the vessel. In a further form, the magneticfield is moved vertically along the height of the reaction vessel todraw the magnetic particles and bound analyte together into a pellet inthe reaction vessel. In a still further form, the moving step moves themagnetic field down from an upper position near the top of the fluid inthe reaction vessel to a position near the bottom of the reaction vesselwhereby the pellet is formed near a bottom side of the reaction vessel.

In yet another aspect of the present invention, a processing zone for aspecimen handling device is provided, including a support adapted tosupport a reaction vessel in a generally vertical orientation, and amagnet supported for selected vertical movement along one side of asupported reaction vessel.

In one form of this aspect of the invention, the support is adapted tosupport a plurality of reaction vessels in at least two rows where eachrow has a defined spacing from at least one adjacent row, and the magnetextends generally horizontally and is supported for selected verticalmovement in the defined spacing. In a further form, the defined spacingbetween rows comprises a generally vertical longitudinal slot, andmagnet is a bar magnet or an electromagnet. In a still further form, (1)the support is adapted to support the plurality of reaction vessels ingenerally parallel first, second, third and fourth rows, (2) the definedspace is a generally vertical longitudinal first slot between the firstand second rows and a generally vertical longitudinal second slotbetween the third and fourth rows, and (3) the bar magnet includes afirst bar magnet supported for selected vertical movement in the firstslot and a second bar magnet supported for selected vertical movement inthe second slot. In a yet further form, the first and second bar magnetsare supported for vertical movement together.

In yet another aspect of the present invention, a method of processinganalytes in fluids in reaction vessels is provided, including the stepsof (1) supporting a reaction vessel containing an analyte in a fluid,(2) binding the analyte to a magnetic particle, (3) introducing amagnetic field into the vessel which draws the magnetic particle andbound analyte to the side of the vessel, and (4) moving the magneticfield vertically along the height of the reaction vessel to draw themagnetic particle and bound analyte together into a pellet in thereaction vessel.

In one form of this aspect of the invention, the moving step moves themagnetic field down from an upper position near the top of the fluid inthe reaction vessel to a position near the bottom of the reaction vesselwhereby the pellet is formed near a bottom side of the reaction vessel.

In a still further aspect of the present invention, a fluid handlingmechanism is provided for an analyte testing device which includes adeck with a processing zone having a plurality of reaction vesselssupported therein with upwardly facing openings. The fluid handlingmechanism includes a first bulk supply of a first fluid, and adispensing head having X discharge openings selectively positionablewith each of the discharge openings over the upwardly facing openings ofX selected reaction vessels, wherein X is at least four. A metering pumpmechanism is adapted to selectively meter X units of a selected quantityof fluid from the first bulk supply, and selectively pump X units ofmetered selected quantities of fluid through the discharge openings tothe selected reaction vessels.

In one form of this aspect of the invention, the deck supports aplurality of reaction vessels in a repeating pattern, each patternincluding X reaction vessels, and the dispensing head discharge openingsare arranged in the pattern. In a further form, the repeating patterncomprises a plurality of rows of reaction vessels. In a still furtherform, the rows include at least eight reaction vessels, and X is atleast eight.

In another form of this aspect of the invention, a second bulk supply ofa second fluid is provided, and the metering pump mechanism is adaptedto selectively meter the X units of the selected quantity of fluid froma selected one of the first and second bulk supplies. In a further form,the metering pump mechanism includes X piston pumps, and X valvestructures are also provided, where each valve structure is associatedwith one of the piston pumps and selectively switchable betweenproviding a connection to (1) a selected one of the first and secondbulk supplies, and (2) an associated discharge opening.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a perspective view of a processing zone and one reactionvessel according to various aspects of the present invention;

FIGS. 2 a, 2 b, and 2 c are cross-sectional views of a processing zonesupporting a plurality of reaction vessels illustrating the magnets indifferent positions according to one aspect of the present invention;

FIG. 3 a is a perspective, cross-sectional view of a processing zonewhich includes the present invention;

FIG. 3 b is an enlarged cross-sectional partial view showing tworeaction vessels supported in the processing zone;

FIG. 4 a is a cross-sectional view of a reaction vessel according to oneaspect of the present invention;

FIGS. 4 b, 4 c, 4 d, and 4 e are enlarged cross-sectional partial viewsof different reaction vessels incorporating one aspect of the presentinvention;

FIG. 4 f is an enlarged cross-sectional partial view of a reactionvessel used with a passive valve as usable with certain aspects of thepresent invention;

FIG. 4 g is an enlarged cross-sectional partial view of a reactionvessel used with an active valve as usable with certain aspects of thepresent invention;

FIG. 5 is a graph showing the theoretical head or height of a particularfluid which will be retained based on the diameter of the fluid bead atthe drain opening;

FIG. 6 is a graph illustrating test results of the head or height of aparticular fluid which will be retained in a reaction vessel havingdifferent hole diameters;

FIG. 7 is a graph illustrating the predicted time required to evacuate a3.5 mL sample through different drain openings; and

FIG. 8 is a diagram of a fluid handling mechanism according to an aspectof the present invention.

DETAILED DESCRIPTION OF THE INVENTION

Two processing zones 20, 22 in accordance with the present invention andusable with a suitable automated testing device (not shown) areillustrated in FIG. 1. For simplicity of explanation of the invention,the overall testing device is not (and need not be) shown in thefigures. For example, a suitable automatic testing device may be adaptedfor the substantial isolation of nucleic acids from biological samples,including the isolation and testing of nucleic acids from biologicalsamples.

In a suitable automated testing device for such a use, for example, ahood may be provided to protect against contamination from theenvironment in which the zones 20, 22 are located to prevent outsidecontaminants from entering therein as is known in the art. Such anautomatic testing device may advantageously also include one or more ofthe following features: (1) a receptacle to hold and segregate fromsamples and reagents used pipette tips such that contamination from usedtips is minimized, (2) aerosol control devices, for example withoutlimitation, (a) a sample tube or reagent tube sealer, (b) electrodes fortreating surfaces and/or liquids with electrical current capable ofmodifying nucleic acids, (c) an ultraviolet light source capable ofdegrading or modifying nucleic acids, (d) an apparatus for causinglaminar air flow in or around the automatic testing device, and (3) anoptical detector (e.g., a flourometer) for measuring the lightabsorbance or fluorescence of processed samples. Tecan AG manufactures ageneral purpose laboratory pipetting instrument with which the variousdescribed aspects of the invention may be used. However, it should beunderstood that many features of such instruments, though usable withthe present invention, do not form a part of the invention, andtherefore are not shown in the figures. Those skilled in the art whoobtain an understanding of the present invention will recognize suchfeatures, such as disclosed in, for example, in U.S. Ser. No. 10/360,956titled “Apparatus and Method for Handling Fluids for Analysis”, filedFeb. 7, 2003, U.S. Pat. No. 6,413,780, titled “Structure and Method forPerforming a Determination of an Item of Interest in a Sample”, and U.S.Publication No. 2002-0127727 also titled “Structure and Method forPerforming a Determination of an Item of Interest in a Sample”, thecomplete disclosures of which are hereby incorporated by reference.

Further, a plurality of processing zones may be used with a singletesting device such as shown, including not only multiple processingzones 20, 22 embodying aspects of the present invention such as shown inFIG. 1, but also additional processing zones (not shown) for differenttypes of processing or specimen handling. For example, additionalhandling zones can be provided wherein reaction vessels may be preparedprior to desired processing by adding specimens, etc. The multipleprocessing zones 20, 22 illustrated in FIG. 1 assist in minimizingresource contention (i.e., conflicts arising where processing using onegroup of reaction vessels may be delayed until another group of reactionvessels has completed processing at the next required processing zone20.

FIG. 1 generally illustrates a processing zone 20 at which testing ofspecimen samples may be done. In suitable testing devices with which theprocessing zones 20, 22 as discussed herein may be used, reactionvessels 26 (only one of which is shown in FIG. 1) containing specimensfor testing may be loaded onto supports 30, 32 at each zone 20, 22. Thesupports 30, 32 may be support brackets or racks, for example, definingrows in which reaction vessels 26 may be supported, each with anupwardly open top. The supports 30, 32 may serve as heat shields toprotect a user from heat blocks therebeneath (as described below).Suitable movable carriers may also be used which are transportable toand from the processing zones 20, 22 (e.g., manually or by a suitablerobotic arm) to facilitate handling reaction vessels where desired.

The supports 30, 32 illustrated in FIG. 1 will support reaction vessels26 in a repeating pattern, with a pattern consisting of a row of eightreaction vessels 26 repeated six times, whereby a total of forty-eightreaction vessels 26 may be processed simultaneously at one processingzone 20 or 22. Thus, in the present description, the processing may beto isolate analytes of interest from up to forty-eight specimens (e.g.,DNA or RNA), after which processing the isolated analyte may be furthertested according to an appropriate protocol. However, it should beunderstood that the present invention is not limited in any way to suchprocessing, and could as readily be used with a device in whichdifferent processing or protocols are carried out.

In the illustrated embodiment, each of the processing zones 20, 22includes heat blocks 40 which may be suitably controlled to heat thereaction vessels 26 to whatever temperatures, for whatever periods oftime, is required by the processing or protocol being carried out. Theheat blocks 40 may be configured so as to surround the reaction vessels26 to dissipate heat from a suitable heater 42 evenly throughout thereaction vessels 26 in the processing zone 20, 22.

The heat blocks 40 may also be arranged with a longitudinal, verticalslot between adjacent rows of reaction vessels 26. For example, asillustrated in FIGS. 2 a-2 c, there is a slot 46 between the left tworows of vessels 26, a slot 48 between the right two rows of vessels 26,and a slot 50 between the middle two rows of vessels 26.

In accordance with one aspect of the invention, suitable bar magnets 60,62, 64 may be supported for movement in the slots 46, 48, 50 between therows of reaction vessels 26. The bar magnets 60, 62, 64 are suitablysupported for selected vertical movement together in the slots 46, 48,50, as by a controlled drive 68 which moves a cross-support 70 onvertical guide rods 72. It would, however, be within the scope of thisaspect of the present invention to use any structure which wouldsuitably control vertical movement of the bar magnets 60, 62, 64 asdescribed hereafter so as to move the magnetic field 76 (see FIG. 2 a)in the reaction vessels 26. Moreover, it would be within the scope ofthe present invention to use still other suitable magnets, such aselectromagnets. Still further, it should be understood that it would bewithin the scope of some aspects of the present invention to not usemagnets at all (e.g., where certain aspects of the invention are used inprocessing which does not magnetically separate the analytes of interestfrom the fluid including, as one example, non-magnetic samplepreparation in which the analytes of interest are bound to silicamembrane).

Specifically, during processing of a specimen within the reaction vessel26, the analyte of interest dispersed in the reaction vessel 26 may besuitably bound to a magnetic material or particles in a suitable mannersuch as is known in the art. During processing thereafter, the magnets60, 62, 64 may be selectively moved vertically in the slots 46, 48, 50to draw the magnetic particles (and bound analyte of interest) to oneside of the vessel 26. Moreover, by selectively moving the magnets 60,62, 64 along the side of the vessels 26 in the slots, the magneticparticles and bound analyte of interest within the reaction vessel 26may be strongly drawn to the side of the vessels 26 throughout theheight of the vessel 26 by essentially subjecting the vessel contents toa uniform magnetic force throughout its height. Further, by moving themagnets 60, 62, 64 from an upper (lyse capture) position such asillustrated in FIG. 2 c to a bottom (wash capture) position asillustrated in FIG. 2 b, magnetic particles and bound analyte ofinterest may not only be drawn to the side of the vessels 26 but mayalso then be pulled down along the side of the vessels 26 as the magnets60, 62, 64 move down whereby a desired pellet of such materials isformed at the bottom corner of the vessels 26.

While the above described structure using bar magnets with multiplevessels 26 may be particularly efficiently used, it should be understoodthat most broadly, aspects of the present invention could also includethe use of a magnet as described with a single reaction vessel 26, thatis, moving a magnet along one side of one vessel 26 to draw the magneticparticles and bound analyte of interest to the side of the vessel 26 anddown to form a pellet at a bottom corner of the vessel 26.

FIGS. 3 a-3 b illustrate yet another aspect of the present invention.Specifically, each processing zone 20, 22 may include concave recesses78 (see FIG. 3 b) for receiving the bottoms of the reaction vessels 26,for example, in the heat blocks 40. A horizontal drain passageway orchannel 80 may extend along the length of the heat blocks 40 beneatheach row of vessels 26, with a vertical passageway 82 connecting thehorizontal passageway 80. A suitable vacuum source (indicatedschematically at 86 in FIG. 3 a) may be applied to the passageways 80,82 to selectively draw/drain fluid from the vessels 26 as describedbelow. While a vacuum source 86 is described herein, it should beunderstood that the significant feature is a lower pressure beneathdischarge or drain openings in the bottom of the vessels 26 (asdescribed in detail below) relative to the pressure on the fluid at thetop of the vessels 26. Therefore, it should be understood that thisaspect of the invention could also be accomplished through theapplication, for example, of high pressure to the top of the reactionvessels 26 where the passageways 80, 82 are at atmospheric pressure, ora combination of pressure and vacuum.

Specifically, as illustrated in FIG. 4 a, it is contemplated as anexample that a reaction vessel 26 may be provided with a fluid 90 andsample having a depth (or height or head) H.

A discharge or drain opening 92 is provided in the bottom of thereaction vessel 26, where the drain opening 92 is configured so that thesurface tension of the fluid 90 in its condition at the processing zone20, 22 is sufficient to support the height of fluid without the fluiddraining through the drain opening 92. It should be understood that,while a single opening 92 is shown in FIG. 4 a, it would advantageouslybe within the scope of the present invention to define the drain openingvia multiple openings through the bottom of the vessel 26.

FIG. 4 b illustrates a reaction vessel 26 with one configuration ofdrain opening 92, where fluid 90 has passed through the drain opening 92so as to form a bead 94 around the opening 92. So long as the opening 92is small enough to maintain a bead 94 which is no greater in size thanthe surface tension of the fluid 90 can maintain, the fluid 90 will besupported in the vessel 26 (that is, until an additional force, arelative pressure between the top and bottom of the fluid 90, isselectively created by the introduction of a vacuum in the passageways80, 82 beneath the vessels 26).

FIG. 4 c discloses an alternative embodiment of a reaction vessel 26 a,wherein a suitable non-wettable coating or surface 96 is provided aroundthe drain opening 92 a. Such a non-wettable coating 96 will prevent abead from spreading out onto the coating (FIG. 4 b illustrates a bead 94which spreads out onto the outer surface of the vessel 26), such that alarger size drain opening 92 a may be used while still maintaining theability of the surface tension of the fluid 90 to support a desiredheight of fluid 90 in the vessel 26 a.

FIG. 4 d discloses still another embodiment of a reaction vessel 26 b,wherein a protruding tube or flange 98 is provided around the drainopening 92 b. The flange 98 may also prevent the bead from spreading outto an area much larger than the flange 98, thereby allowing use of alarger drain opening 92 b while still maintaining the ability of thesurface tension of the fluid 90 to support the fluid 90 as described forFIG. 4 c.

FIG. 4 e discloses yet another embodiment of a reaction vessel 26 c, inwhich a hydrophobic frit 99 or other suitable hydrophobic porousmaterial (such as may be obtained from Porex Corporation of Fairburn,Ga.) is associated with the vessel 26 c to define the drain opening 92c. The hydrophobic frit 99 may be advantageously selected, based on thefluid, whereby the frit 99 will support the desired height of fluid 90in vessel 26 c, and will allow the fluid to pass therethrough when aselected pressure differential is introduced between the top of thevessel 26 c and beneath the frit 99/drain opening 92 c. Thus, where afluid having low surface tension properties is used (e.g., alcohol), theporosity of the frit material may advantageously be less than thematerial used with fluids having higher surface tension properties toenable the desired height of fluid to be supported as desired.

The above described vessels 26, 26 a, 26 b, 26 c are particularlyadvantageous inasmuch as such vessels are low cost disposables. However,it should be understood that aspects of the present invention encompassstill further vessels having drain openings which will support a head offluid 90 by other than the fluid surface tension, while allowing thatfluid to be selectively drained from the vessel responsive to a selectedpressure differential created (e.g., by the introduction of a vacuum inthe passageways 80, 82 beneath the vessels 26).

For example, a drain opening consisting of not only an opening in thevessel 26 but also a suitable passive valve may be provided to providethe desired fluid flow, where the passive valve is biased to block fluidflow unless a selected pressure differential across the valve iscreated. FIG. 4 f illustrates a reaction vessel 26 d having a duckbillvalve 100 as one example of a suitable valve which would be pulled openby a selected pressure differential resulting from a vacuum in thepassageways 80, 82. Those skilled in the art will recognize that manyother valve structures providing such operation would be suitable,including, for example, umbrella valves, flapper valves and springbiased ball check valves, and could also be advantageously used withcertain aspects of the present invention.

Active valves may also be advantageously used with certain aspects ofthe present invention. FIG. 4 g illustrates a reaction vessel 26 e witha suitable connection 102 to passageways 80′, 82′ with a suitable pinchvalve 104 and suitable control 106 for selectively closing and openingthe passageway 80′ as desired for operation. A hydrophobic frit 99′ mayalso be provided in the bottom opening of the vessel 26 e. Active valvesmay advantageously be selected which will support the desired height offluid and may be opened to allow fluid to drain from the vessel 26 ewithout assist by a pressure differential. However, it would be withinthe scope of certain aspects of the present invention to additionallyprovide a vacuum to assist in such draining when the active valves 104are open.

Valves such as described above may be a part of the reaction vessel, orpart of the vacuum passageways 80, 82 beneath the vessel.

FIG. 5 illustrates a theoretical retention height H of the followingfluid 90, such as may be commonly encountered as one example:

Vessel Content 1 mL Plasma + 2.5 mL Buffer Solution Sample height 5 cmTemperature 50 degrees C. Sample density ρ 1 g/cc Sample surface tensionσ 60 dyne/cm (est. at 50 degrees C.) Sample viscosity μ 0.87 cP (est. at50 degrees C.)As can be seen from the curve 106 in FIG. 5, for such a fluid, the 60dyne/cm surface tension would be sufficient to support a height H offluid of 5 cm if the bead diameter is slightly less than 0.02 inches (orsmaller). Thus, any drain opening 92 which will form a bead diameter nolarger than slightly less than 0.02 inches will be able to retain thefluid 90 in the reaction vessel 26 until a relative pressure isintroduced via the vacuum 86.

As illustrated in FIGS. 4 b and 6, however, the drain opening 92 as inFIG. 4 b may not be as large as the allowable bead size inasmuch as thebead may tend to spread out around the opening 92. FIG. 6 shows theresults 110, 112, 114 of tests using DI water at 22 degrees C. (with asurface tension a of 72.7 dyne/cm), in which it can be seen that theactual height of supported liquid 90 (generally around 1 to 2 cm) for agiven hole diameter was far below the maximum theoretical liquidretention height 118 for a bead of such water having that same diameter.Such a shortfall can be attributed in large part to the spreading of thebead (see 94 in FIG. 4 b) around the opening 92. As previouslydiscussed, the FIGS. 4 c and 4 d embodiments address this issue bylimiting the spreading of the bead around the opening 92 a, 92 b.

FIG. 7 illustrates the predicted time to evacuate a 3.5 mL sample (e.g.,a 5 cm fluid height from a conventional reaction vessel 26) based on thehole diameter. Different theoretical conditions are illustrated usingthe FIG. 5 sample fluid (having a fluid viscosity μ of 0.87 cP at atemperature of 50 degrees C.). Hole lengths of 0.040 inch and 0.080inch, and vacuum of −10 inches Hg and −20 inches Hg, are illustrated.Specifically, the theoretical evacuation time for a hole length of 0.080inch using a −10 inch Hg vacuum is shown at 120, the theoreticalevacuation time for a hole length of 0.040 inch using a −10 inch Hgvacuum is shown at 122, the theoretical evacuation time for a holelength of 0.080 inch using a −20 inch Hg vacuum is shown at 124, and thetheoretical evacuation time for a hole length of 0.040 inch using a −20inch Hg vacuum is shown at 126.

As can be seen from the FIG. 7 curves 120, 122, 124, 126, the holelength has a theoretical minor impact on the predicted evacuation time.The amount of vacuum assist has a greater impact on the time, but thatis still relatively small. Of particular significance is the indicationthat hole diameters which are about 0.01 inch or smaller will havesignificantly greater evacuation times, with holes much smaller thanabout 0.005 inch theoretically incapable of evacuating the fluid withthe indicated vacuum levels. Since evacuation time could thuspotentially significantly slow processing of samples, it should beappreciated that the larger hole diameters above 0.01 inch would provideadvantageous speeding of processing over smaller hole diameters. Thus,the FIGS. 4 c and 4 d embodiments, for example, which will enable thereaction vessels 26 a, 26 b to retain the desired height of fluid 90with larger size drain openings 92 a, 92 b such as previously described,may be particularly advantageously used with the present invention.

This manner of draining fluid 90 from the reaction vessels 26 shouldthus be appreciated to be fast and convenient. Further, it should beappreciated that such draining may be accomplished with minimal cost ofdisposable pipettes. Moreover, it should be appreciated that the use ofreaction vessels 26 with bottom drain openings 92 such as described maybe advantageously used with the previously described movable magnets 60,62, 64, inasmuch as the magnets 60, 62, 64 operate to pull the magneticparticles and bound analyte of interest to the bottom side of the vessel26, whereby the pellet of such material will be clear of the drainopening 92.

FIG. 8 discloses a further aspect of the present invention, which uses atraveling head 200 connected to bulk supplies of fluid, whereby desiredamounts of such fluids may be added to sets of vessels 26 (not shown inFIG. 8).

Specifically, the head 200 includes two sets of outlets 204, 206, withone outlet set 204, for example, used for wash, and a second outlet set206 used for wash and pipette prime. A second (or additional) outlet set206 may be provided, for example, where a different type of discharge(e.g., a spray nozzle) may be desired. In the illustrated embodiment,the outlet sets 204, 206 include eight outlets arranged in a row tomatch the pattern of the vessels 26 supported in the processing zones20, 22. Thus, the head 200 may be arranged above any selected patternrow of eight vessels 26 in a processing zone 20, 22 whereby the eightoutlets of a selected set 204, 206 will be aligned above the selectedeight vessels 26 so that fluid discharged from the head outlets willenter the selected vessels 26.

Associated with the head 200 is a suitable pump 220 which may meterdesired amounts of selected fluids (as further described below) for eachof the outlets in a set 204, 206. One such suitable metering pump 220 isillustrated in FIG. 8 as a Cavro 24V, 48V signal motor which includeseight 2.5 mL piston pumps 222. Cavro Scientific Instruments Inc. islocated at 2450 Zanker Road, San Jose, Calif., USA 95131. As describedfurther herein, this pump 220 will meter a desired amount (e.g., 2.5 mL)of fluid from the bulk supplies for each of the X number (eight in theillustrated embodiment) outlets of each outlet set 204, 206. However, itshould be understood that the details of this pump 220 do not form apart of the invention, and any pump and valving system which will metera selected number (X, e.g., eight) of a selected quantity (e.g., 2.5 mL)of fluid for discharge through the outlets of a selected outlet set 204,206 would be suitable.

Suitable bulk supplies 230, 232 may be provided according to theexpected needs of the testing. In the illustrated example, there is abulk supply 230 of wash and a bulk supply 230 of single step lysisbuffer (SSLB). As illustrated in FIG. 8, each bulk supply 230, 232 mayinclude a refillable tank 236, 238 which is connected to a sealeddispensing tank 240, 242. A valve 246, 248 may selectively connect thedispensing tank 240, 242 to a vacuum source (vacuum reservoir 254 andvacuum pump 256) to assist in maintaining a desired level of fluid inthe dispensing tank 236, 238, and to permit fluid to be drawn off thetop of the dispensing tanks 240, 242 if desired. Another vacuum valve260 may be used to selectively draw such materials to a waste container262. It should be understood, however, that the illustrated bulk supplystructure is merely one suitable example of a structure which may beused with this aspect of the invention, and any suitable bulk supplyfrom which the needed fluids may be pumped by the metering pump 220 maybe used with this aspect of the present invention.

The dispensing tanks 236, 238 are suitably connected to the travelinghead 200, as by flexible hoses 270, 272.

A suitable valve structure is provided to enable the metering pump 220to be selectively connected to the bulk supply of selected fluid inorder to obtain X (e.g., eight) units of selected quantity (e.g., 2.5mL), after which the X units of selected fluid may be sent to a selectedset of outlets 204, 206 for discharge into a selected set of reactionvessels 26 over which the head 200 has positioned the selected outletset 204, 206.

One valve structure which would be suitable for a head 200 connected totwo bulk supplies 236, 238 and having two outlet sets 204, 206 is thethree-valve structure illustrated in FIG. 8. One such valve structure isassociated with each of the piston pumps 222 illustrated. While theillustrated embodiment may be advantageously used with this aspect ofthe invention, it should be recognized that this aspect of the inventionmay be readily practiced with different valve structures.

Specifically, the illustrated three-valve structure includes valves 280,282, 284, each of which may be selectively switched between path A andpath B. During a single cycle, for example, valve 280 may be connectedto path A, after which the metering pump 220 may be activated to draw2.5 mL of wash fluid from bulk supply 230 through hose 270 into thepiston pumps 222. Valve 280 may then be switched to path B, valve 282switched to path A, and valve 284 switched to path B, whereby the pistonpumps 222 may then be operated to discharge the eight 2.5 mL units ofwash fluid through the eight outlets of outlet set 284 into vessels 26(not shown) located beneath those outlets.

When used with a processing zone 20, 22 in which there are six rows ofeight reaction vessels 26 such as previously described, the aboveprocess may be repeated six times to provide the wash fluid to allforty-eight reaction vessels 26.

After the wash fluid has been discharged into all of the selectedreaction vessels 26, operation of the valve structure can be changed tosupply a different fluid if needed based on the testing beingaccomplished. For example, if SSLB fluid is thereafter desired, valve280 positioned at path B, valve 282 positioned at path A, and valve 284positioned at path A, whereby the piston pumps 222 may then be operatedto draw 2.5 mL of SSLB fluid from bulk supply 232 through hose 272 intothe piston pumps 222. Then, valve 280 may be kept at path B and valve282 switched to path B, whereby the piston pumps 222 may then beoperated to discharge the eight 2.5 mL units of SSLB fluid through theeight outlets of outlet set 284 into vessels 26 (not shown) locatedbeneath those outlets. This processing may then be repeated as necessaryto provide SSLB fluid to all of the selected reaction vessels 26.

It should be appreciated that the FIG. 8 aspect of the invention willenable the processing zones 20, 22 to be used efficiently and reliably.The desired amounts of fluid may be easily and reliably metered in thedesired amounts. Further, this may be accomplished quickly, without thedelay time which would be required by a dispensing head which travelsback and forth from the processing zones and bulk supplies each time aset of reaction vessels requires such fluids.

It should also be appreciated that the various aspects of the inventiondescribed herein may be combined to provide a processing zone which maybe advantageously operated to efficiently and quickly process samples.

Still other aspects, objects, and advantages of the present inventioncan be obtained from a study of the specification, the drawings, and theappended claims. It should be understood, however, that the presentinvention could be used in alternate forms where less than all of theobjects and advantages of the present invention and preferred embodimentas described above would be obtained.

1. A method of processing analytes in fluids in reaction vessels,comprising: supporting a reaction vessel containing an analyte in afluid, said reaction vessel having a drain opening in its bottom capableof supporting a selected head of said fluid in said vessel by thesurface tension of the fluid; drawing said analyte to a side of saidvessel to concentrate said analyte clear of said drain opening; andselectively creating a pressure differential between the top of thefluid and the bottom of the drain opening sufficient to overcome saidfluid surface tension and drain the fluid through the drain 12 opening.2. The method of claim 1, wherein said selectively creating a pressuredifferential step comprises selectively creating a vacuum beneath saiddrain opening.
 3. The method of claim 1, wherein said drawing stepcomprises: binding said analyte to a magnetic particle, and introducinga magnetic field into said vessel which draws the magnetic particle andbound analyte to the side of the vessel.
 4. The method of claim 3,further comprising moving said magnetic field vertically along theheight of said reaction vessel to draw said magnetic particles and boundanalyte together into a pellet in said reaction vessel.
 5. The method ofclaim 4, wherein said moving step moves said magnetic field down from anupper position near the top of the fluid in the reaction vessel to aposition near the bottom of the reaction vessel whereby said pellet isformed near a bottom side of said reaction vessel.
 6. A processing zonefor a specimen handling device, said processing zone comprising: asupport adapted to support a reaction vessel in a generally verticalorientation; and a magnet supported for selected vertical movement alongone side of a supported reaction vessel.
 7. The processing zone of claim6, wherein: said support is adapted to support a plurality of reactionvessels in at least two rows where each row has a defined spacing fromat least one adjacent row; and said magnet extends generallyhorizontally and is supported for selected vertical movement in saiddefined spacing.
 8. The processing zone of claim 7, wherein said definedspacing between rows comprises a generally vertical longitudinal slot,and said magnet is an electromagnet.
 9. The processing zone of claim 7,wherein said defined spacing between rows comprises a generally verticallongitudinal slot, and said magnet is a bar magnet.
 10. The processingzone of claim 9, wherein: said support is adapted to support theplurality of reaction vessels in generally parallel first, second, thirdand fourth rows; said defined space comprises a generally verticallongitudinal first slot between the first and second rows and agenerally vertical longitudinal second slot between the third and fourthrows; and said bar magnet comprises a first bar magnet supported forselected vertical movement in the first slot and a second bar magnetsupported for selected vertical movement in the second slot.
 11. Theprocessing zone of claim 10, wherein said first and second bar magnetsare supported for vertical movement together.
 12. A method of processinganalytes in fluids in reaction vessels, comprising: supporting areaction vessel containing an analyte in a fluid; binding said analyteto a magnetic particle; introducing a magnetic field into said vesselwhich draws the magnetic particle and bound analyte to the side of thevessel; and moving said magnetic field vertically along the height ofsaid reaction vessel to draw said magnetic particle and bound analytetogether into a pellet in said reaction vessel.
 13. The method of claim12, wherein said moving step moves said magnetic field down from anupper position near the top of the fluid in the reaction vessel to aposition near the bottom of the reaction vessel whereby said pellet isformed near a bottom side of said reaction vessel.
 14. A fluid handlingmechanism for an analyte testing device, said testing device including adeck with a processing zone having a plurality of reaction vesselssupported therein with upwardly facing openings, said fluid handlingmechanism comprising: a first bulk supply of a first fluid; a dispensinghead having X discharge openings selectively positionable with each ofsaid discharge openings over said upwardly facing openings of X selectedreaction vessels, wherein X is at least four; a metering pump mechanismadapted to selectively meter X units of a selected quantity of fluidfrom said first bulk supply, and selectively pump said X units ofmetered selected quantities of fluid through said discharge openings tosaid selected reaction vessels.
 15. The fluid handling mechanism ofclaim 14, wherein: said deck supports a plurality of reaction vessels ina repeating pattern, each pattern including X reaction vessels; and saiddispensing head discharge openings are arranged in said pattern.
 16. Thefluid handling mechanism of claim 15, wherein said repeating patterncomprises a plurality of rows of reaction vessels.
 17. The fluidhandling mechanism of claim 16, wherein X is at least eight and saidrows include at least eight reaction vessels.
 18. The fluid handlingmechanism of claim 14, further comprising a second bulk supply of asecond fluid, wherein said metering pump mechanism is adapted toselectively meter said X units of said selected quantity of fluid from aselected one of said first and second bulk supplies.
 19. The fluidhandling mechanism of claim 18, wherein said metering pump mechanismincludes X piston pumps, and further comprising X valve structures, eachvalve structure being associated with one of said piston pumps andselectively switchable between providing a connection to (1) a selectedone of said first and second bulk supplies, and (2) an associateddischarge opening.